Flow cytometry is a laser-based, biophysical technology where fluorescent molecules coupled to cells are aspirated into the flow cytometer, then passed one by one through a flow cell and excited by a set of lasers. The emitted fluorescence is then collected and transformed into an electrical signal for analysis. Labelling cells with molecules that fluoresce at different wavelengths allows the user to identify a variety of distinct cell populations and therefore it provides a powerful tool with diagnostic, therapeutic, and research applications. For example, the technique is often used to count cells from a biological sample, such as counting CD4/CD8 populations from circulating blood in human immunodeficiency virus (HIV) studies to map the disease, and to sort cells from a mixed population of cells, such as stem cell sorting from harvested biological samples for possible differentiation and reintroduction into different areas of the patient's body.
Since numerous distinct cell populations can be identified using flow cytometry often a single flow cytometry experiment or test includes a large number of samples. Therefore, there is a need to increase the throughput of flow cytometers. One solution is to present test samples on multi-well plates and program a robot to acquire samples across the multi-well plates, which is conventionally performed by aspirating cell samples from the wells. However, a challenge with presenting cells in multiwall plates is that cells tend to settle from solution and collect at the bottom of wells while waiting for aspiration, thereby decreasing cell yields in later aspirated samples.
A possible solution to the problem of cell settling is to rotate or agitate multi-well plates at various intervals to assist in suspending cells for aspiration. We attempted different intervals including between steps of aspirating samples from different wells. However, it has now been found that merely using a conventional suspension approach of rotating or agitating the multi-well plate, even at frequent intervals, only significantly helps suspend cells positioned at opposing ends of the plate. Importantly, it is found to be insufficient to suspend cells positioned within the middle region of the plates. That is, the approach tends to suspend cells in the outer region of the multi-well plate but fails to adequately suspend cells at the middle region of the plate. Accordingly, there is a need to develop an approach useful in high throughput sample acquisition for flow cytometry procedures that can increase the consistency of cell yields across the entire multiwall plate.